Structural Studies of the Vitamin D Receptor

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DESCRIPTION (provided by applicant): The steroid and nuclear hormone receptors are the largest known class of eukaryotic transcriptional regulators and modulate transcription in response to non-peptide lipophillic signals. The members of this superfamily include the vitamin D receptor (VDR), which regulates genes related to calcium homeostasis and plays a role in the differentiation of hematopoietic and skin cells, and the androgen receptor (AR), which directly regulates the expression of prostate-specific genes, among others. Hormone receptors exert their effects by binding as homo- or hetero-dimers to DNA targets - called response elements - composed of two hexanucleotide half-sites that are distinguished from one another by the sequence of the half site and the orientation and spacing of one half site relative to the other. VDR binds to response elements composed of two half-sites arranged as a direct repeat with three base pairs of spacer in between (DR-3). AR binds to inverted repeat, three-spacer response elements (IR-3), although recently a novel AR response element was identified that closely resembles the DR-3 type and may serve to distinguish AR responsive genes from those responsive to glucocorticoids and other steroids. An understanding of the underlying stereochemistry of the role of VDR and AR in transcriptional activation and DNA target selection requires the structural analysis of the relevant macromolecular species and complexes. We plan to solve and analyze, using X-ray crystallography, the structures of the DNA binding domain of VDR homodimers and RXR-VDR heterodimers in complex with a variety of DNA targets, and the structure of the AR DNA binding domain in complex with its novel direct repeat DNA target. These studies are aimed at elucidating the structural basis for both VDR and AR DNA target recognition, their shared preference for DR-3 type response elements, and the mechanisms employed to distinguish correct from incorrect DNA targets. We will correlate our structural results with measurements of the protein's binding affinity to consensus and wild-type response elements. NMR and crystallography will be used in parallel and synergistically to examine the structure of full-length VDR, in order to understand the interplay between the DNA and ligand binding domains. These analyses may lead to the design of novel anti-cancer or therapeutic compounds.